Related Experiment Video
Updated: Aug 9, 2025
Improved Enzyme Protection Assay to Study Staphylococcus aureus Internalization and Intracellular Efficacy of Antimicrobial Compounds
Published on: September 8, 2021
Non-Specific Signal Peptidase Processing of Extracellular Proteins in Staphylococcus aureus N315
Santosh A Misal1, Shital D Ovhal1, Sujun Li2
1Department of Chemistry, Indiana University, 800 E Kirkwood Avenue, Bloomington, IN 47405, USA.
Abstract:
Staphylococcus aureus is one of the major community-acquired human pathogens, with growing multidrug-resistance, leading to a major threat of more prevalent infections to humans. A variety of virulence factors and toxic proteins are secreted during infection via the general secretory (Sec) pathway, which requires an N-terminal signal peptide to be cleaved from the N-terminus of the protein. This N-terminal signal peptide is recognized and processed by a type I signal peptidase (SPase). SPase-mediated signal peptide processing is the crucial step in the pathogenicity of S. aureus. In the present study, the SPase-mediated N-terminal protein processing and their cleavage specificity were evaluated using a combination of N-terminal amidination bottom-up and top-down proteomics-based mass spectrometry approaches. Secretory proteins were found to be cleaved by SPase, specifically and non-specifically, on both sides of the normal SPase cleavage site. The non-specific cleavages occur at the relatively smaller residues that are present next to the -1, +1, and +2 locations from the original SPase cleavage site to a lesser extent. Additional random cleavages at the middle and near the C-terminus of some protein sequences were also observed. This additional processing could be a part of some stress conditions and unknown signal peptidase mechanisms.
Insights
This study reveals that Staphylococcus aureus signal peptidase (SPase) exhibits both specific and non-specific cleavage of secreted proteins. These findings uncover novel processing mechanisms beyond the canonical SPase cleavage site.
Area of Science:
- Microbiology
- Proteomics
- Molecular Biology
Background:
- Staphylococcus aureus is a significant human pathogen with increasing multidrug resistance.
- Secretion of virulence factors via the general secretory (Sec) pathway is crucial for S. aureus pathogenicity.
- Type I signal peptidase (SPase) processes N-terminal signal peptides, a critical step in protein secretion.
Purpose of the Study:
- To investigate the cleavage specificity of SPase in S. aureus.
- To identify novel N-terminal protein processing events mediated by SPase.
- To understand the implications of SPase activity on S. aureus pathogenicity.
Main Methods:
- Employed a combination of N-terminal amidination, bottom-up, and top-down proteomics.
- Utilized mass spectrometry-based approaches to analyze protein processing.
- Evaluated SPase-mediated cleavage sites on secretory proteins.
Main Results:
- SPase demonstrated both specific and non-specific cleavage of secretory proteins.
- Non-specific cleavages were observed adjacent to the canonical SPase cleavage site (-1, +1, +2 positions).
- Additional random cleavages occurred in the middle and near the C-terminus of some proteins.
Conclusions:
- SPase exhibits broader cleavage activity than previously understood.
- Unusual cleavage patterns may be linked to stress conditions or uncharacterized SPase mechanisms.
- Further research is needed to elucidate the biological significance of these novel processing events.
Related Concept Videos
01:28Peptidoglycan Synthesis
01:26Bacterial Protein Maturation
01:39Mitochondrial Precursor Proteins
Most of the mitochondrial...